Addition compounds of amino acids and hydrofluoric acid or soluble fluorides,and method of preparing the same



United States Patent 20 Claims. cl. 260429.3)

ABSTRACT OF THE DISCLOSURE Addition compounds of lower aliphatic aminoacids with hydrogen fluoride or soluble metal fluoride are effective inthe prophylactic treatment of caries. The novel compounds are preparedby reacting amino acid with HF, and recovering the addition compoundfrom the reaction mixture. It is also possible to hydrolyze analbuminous substance such as egg albumen, wheat gluten or the like byheating with cone. HF, neutralizing the reaction mixture with metalhydroxide, and evaporating to dryness.

This application is a division of U.S. application Ser. No. 361,245,filed Apr. 20, 1964, now abandoned.

This invention relates to addition compounds of amino acids withhydrogen fluoride or fluoride salts, and to a method for theirpreparation.

Research during recent years has shown that the element fluorine playsan important part in the constitution of the hard dental substance andthat the lack of this element leads to diminished resistance to caries.It is :also known that the artificial introduction of this constituentin careful dosage brings about a significant reduction in the incidenceof caries. In view of the fact that over 90% of the population of thecivilized world suffer to a greater or lesser degree from dental cariesthe recognition of the :above mentioned fact is of a social and hygienicimportance that cannot be underestimated. The supplementary introductionof the element fluorine has therefore been widely discussed in recentyears and a therapy consisting in the provision of artificiallyfluori-nated food components, preferably in the form of additives todrinking water, salt or milk, as well as in the form of tablets has beenactively practiced.

It has also been shown that once a tooth has erupted the above mentionedmeasure cease to be effective or at least that their eflicacy is greatlyreduced, but that the tooth is able to absorb fluorine ions from theoutside, to bind them chemically, and thence to improve its resistanceto caries. This fact gave rise to the prophylactic treatment of cariesby topical application of fluoride solutions to the teeth. Although suchtreatment provides an excellent measure of protection, it is timeconsuming and expensive and, thus, is not always available to peopledesiring it. The most practical means of administering fluorine lies inthe use of dentifrice or chewing gum compositions.

According to the prior art, fluorine was used substantially in the formof its metal salts such as stannous or sodium fluoride. It has now beenfound that certain addition compounds of soluble fluorides with loweraliphatic amino acids provide an even greater degree of anticariogenieactivity than has been obtained with the metal fluorides.

Accordingly, it is an object of this invention to provide novel fluorinecompounds having improved anticariogenic activity.

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A further object is to provide a novel method for preparing these newcompounds.

Other objects and advantageous features will be obvious from thefollowing detailed description.

Unexpectedly, addition compounds of lower aliphatic amino acids withhydrogen fluoride or soluble metal fluorides have been found to beeffective in the prophylactic treatment of caries, many of thesecompounds having an effect surpassing that of the forementioned metalsalts. These compounds are addition compounds having the general formulaRX where R is a lower aliphatic amino acid and X is a soluble fluoridewhich normally dissociates to yield fluoride ions in aqueous solution.Preferably, the lower aliphatic amino acids will contain not more thanabout 10 carbon atoms in the aliphatic chain. Suitable amino acidsinclude the monoaminomonocarboxylic acids, such as carbamic acid,glycine, alanine, valine, norvaline, leucine, norcleucine, isoleucine,isovaline, phenylalanine, tyrosine, serine,alpha-amino-beta-hydroxybutyric acid, cysteine, cystine, betaine andsarcosine. Monoamino-dicarboxylic acids such as aspartic and glutamicacid can also be employed. Operable diamino-monocarboxylic acids includearginine, lysine, ornithine, asparagine and citrulline.

Soluble fluorides which normally dissociate in aqueous solution to yieldfluoride ions and can thus comprise the fluoride portion of the additioncompounds of this invention include, for example, HF, SnF KF, and ZrFHydrofluoric acid and stannous fluoride are especially preferred solublefluorides. The addition product of an amino :acid with HF is an aminoacid hydrofluoride. The addition product may be described as a doublesalt when the soluble fluoride portion is a metal salt.

Amino acid hydrofluorides can be prepared in accordance with thisinvention by reacting amino acids, either in the dry state or insolution or suspension in suitable media (as more fully hereinafterexplained), with aqueous hydrofluoric acid or gaseous hydrogen fluoride.The solvent is then partially evaporated and the hydrofluoride isallowed to crystallize from the cooling mother liquor from which it isthen separated. In the case of hydrofluoride salts that do not readilycrystallize, the solvent is completely evaporated and the salts remainas a dry or syrupy residue. If left for several days over concentratedsulfuric acid with the exclusion of air, the syrup can be made tocrystallize. However, if the hydrofluoride occurs in the form of asuspension it may be separated by filtration. The fluoride portion ofthe addition compound thus prepared will consist of at least about 1 molof hydrofluoric acid.

The addition compounds of amino acids and soluble fluorides can beprepared in a simple manner by directly reacting the components in asuitable solvent, for example water. The compounds may be crystallizedin concentrated solutions or they may be obtained by evaporation of thesolution to dryness.

The invention will be illustrated by reference to a number of exampleswhich are, however, not intended to limit the scope of the invention inany way.

Example I 21.5 g. of glycine together with 19.75 g. of a 38%hydrofluoric acid are heated in a platinum dish over a Water bath. Thewater is then evaporated from the clear solution. By adding 10milliliters of ethyl alcohol and evaporating again more of the water canbe removed. In the course of cooling, the hydrofluoride separates in theform of colorless crystals.

The substance is difficultly soluble in methyl and ethyl alcohol,insoluble in acetone, readily soluble in water.

Formula: HF.NH CH COOH 3 Example II 17.8 g. of sarcosine and 22.2 g. ofa 60% hydrofluoric acid are mixed in a platinum dish. The sarcosinedissolves with the evolution of heat. The dihydrofluoride formed isallowed to crystallize in a sulfuric acid desiccator. The crystals arethen quickly filtered off by suction, remoistened with about 5 ml. ofwater, and filtered off again by suction. The product is dried in acurrent of warm air between 40 and 50 C.

The substance consists of colorless crystals which are soluble in hotmethyl alcohol, difliculty soluble in ethyl alcohol, insoluble inacetone, and easily soluble in water.

The formula of the compound is:

Example III 21.9 g. of lysine are dissolved in 75 cc. of water and 9.2g. of a 38% hydrofluoric acid added to the solution. The reactionmixture is evaporated to dryness on a water bath and the residue dried.

The crystals are colorless, dilficulty soluble in methyl and ethylalcohol, insoluble in acetone and readily soluble in water.

Formula:

Example IV 17.8 g. of alanine are dissolved in 21.0 g. of a 38%hydrofluoric acid and evaporated on a water bath. Evaporation is twicerepeated, 20 cc. of ethyl alcohol being added each time, and the residueis then dried in vacuo at 50 to 60 C. for two hours. Upon cooling, thesyrupy residue gradually begins to crystallize. After separating thecrystals by sharp filtration by suction, they are moistened with about 5ml. of water and again filtered by suction. The product is thencarefully dried in a warm current of air at about 40 C.

Alanine hydrofluoride crystallizes in colorless prisms that aredifiicultly soluble in ethyl alcohol, slightly more soluble in methylalcohol, and easily soluble in Water.

The compound has the following formula:

011 01100 OILHF Example V 11.6 g. of potassium fluoride are dissolved in30 cc. of water and 23.4 g. of butaine are added to the solution. Theclear solution is partly evaporated on a water bath and then allowed tocrystallize. The crystal pulp is filtered by suction, washed with about5 cc. of alcohol and dried.

The compound consists of colorless crystals, soluble in methyl alcohol,slightly less in ethyl alcohol, but easily soluble in water.

Formula:

Example VI 16.7 g. or zirconium fluoride are dissolved in 150 cc. ofwater with the simultaneous addition of 4 drops of a 38% hydrofluoricacid. 11.7 g. of betaine are then added to the solution.

The clear reaction mixture is evaporated on a water bath untilcrystallization begins. It is then cooled and the crystalline mass israpidly filtered off by suction. The product is dried in vacuo at 50 to60 C.

The compound consists of colorless crystals, which are diflicultysoluble in ethyl and methyl alcohol, insoluble in acetone, and easilysoluble in water.

Formula:

4 Example VII 15.7 g. of stannous fluoride are dissolved in cc. of hotwater with the simultaneous addition of 4 drops of a 38% hydrofluoricacid, 7.5 g. of glycine being likewise added to the solution. The clearreaction mixture is evaporated over a Water bath until crystallizationbegins, then cooled, and the crystalline mass quickly filtered off bysuction. The product is dried in vacuo at 50 to 60 C.

15.7 g. of stannous fluoride with the addition if necessary, of 1 to 4drops of a 38% hydrofluoric acid are dissolved in cc. of hot water and7.5 g. of glycine are added thereto. The solution is evaporated until ithas the consistency of a syrup, and then allowed to crystallize. At theend of one or two days the compound has fully crystallized. The crystalsare colorless, difliculty soluble in ethyl and methyl alcohol, insolublein acetone, and readily soluble in water. The aqueous solution easilyhydrolyzes, forming the basic stannous fluoride. By adding one drop ofhydrofluoric acid hydrolysis can be inhibited.

Formula: NH CH COOHSnF Example VIII Aspartic acid is reacted withstannous fluoride in the manner described in Example VII.

Formula:

NIIzCHCHgCOOILSllFg O OH Each of the following addition compounds can beprepared by reacting the indicated amino acid with soluble fluorides:

betaine liydrotluorido (C113) 31}! Cl'lgU O 0.111"

NI-I

CH3CHCOOH.S11F;

NH CHgCOOIIKF oHiNHc-HmooHKF OH CHQCHZoHCOOHsnFZ N11 glutamic acidstannous fluoride NHQGILC O OH.SnF

CHzCHzCOOH CHKNHCH'JCOOH.ZIF4 In like manner all of the amino acidsdescribed as operable herein can be reacted with the above salts orhydrofluoric acid to yield the corresponding addition product.

Mixtures of amino-acid hydrofluorides can be conveniently prepared byhydrolysis of albuminous substances with hydrofluoric acid. Thesehydrolysates can be obtained by heating albuminous substances, such asegg albumen, blood albumen, casein, gluten, keratin, for several hourswith concentrated hydrofluoric acid (preferably to The reaction time canvary substantially, hydrolysis and addition being sufliciently completefor the purposes of this invention in about 7 hours. Longer reactiontimes can be employed but there is no substantial improvement in yield.After hydrolysis the reaction mixture is evaporated to dryness. Theresidues contain an average of 1020% HF and are very largelywater-soluble. Preferably the mixture is neutralized with metalhydroxides before final evaporation. The product then consists ofaddition compounds of fluorides with the amino acids formed, as well asof free fluorides. The metal hycysteine hydrotlnoride sarcosine stannousfluoride lysine stannous fluoride alanine stannous fluoride glycinepotassium fluoride sarcosine potassium fluoride norvaline stannousfluoride sarcosine zirconium fluoride droxides employed in theneutralization step are those which form soluble fluorides, e.g., KOH,NaOH, Sn(OH) and LiOH.

Example IX 15 g. of wheat gluten are heated with 60 g. of 38%hydrofluoric acid on a water bath for 7 hours and then evaporated todryness in vacuo at 60 C.

17 g. of a brown residue which is nearly wholly soluble in water andcontaining 17.0% HF remains. The residue consists predominantly of amixture of hydrofluorides of the amino acids arginine, lysine, tyrosine,phenylalanine, cystine, methionine, threonine, leucine, isoleucine,valine, glycine, alanine and glutamic acid.

Example X The anticariogenic eflicacy of the compounds of this inventionwas established in the following manner. Freshly extracted, mainlycaries-free molars and bicuspids which were stored in physiologic salinefrom 1 to days at a temperature of 2 C., were mechanically cleaned andbrushed with tap water and pumice. The roots and occasional initialcaries lesions were covered with wax. The crowns were then exposed for420 hours to the substance to be tested, in aqueous solutions (50milliliters) at a pH of 2.9. After this treatment, the teeth were washedfor 30 minutes in physiologic saline, the latter being agitated andrenewed three times. The crowns were not brushed. They were decalcifiedin a horizontal shaker at a temperature of 37 C. in 12.5 cc. of aphthalate buffer and distilled water at a 1:1 ratio and at a pH of 4.The protected enamel of the teeth was demineralized during a three hourperiod (3 hour values). The following values were obtained when treatingteeth in the above defined way with the indicated compounds. In thefollowing table the designation S.R.R. refers to the solution ratereduction in percent for calcium and phosphorus removal from intactenamel treated with the indicated compounds at a pH of 2.9 and afluorine concentration of 0.1%, the treatments having been carried outfor three hours as stated above.

Table Compound tested: S.R.R. percent (a) Sodium fluoride 71 (b)Stannous fluoride 69 (c) Lysine-KP 87 (d) Leucine'HF 84 (e) Cystine-HF79 (f) Cystine amine-HF 92 It will be observed that the values obtainedemploying representative amino acid compounds of this invention whencompared with sodium fluoride and stannous fluoride, both of whichsubstances are considered to be highly active against dental cariesformation, the former compounds are manifestly superior. It is assumedthat the improved enamel protection is arrived at by the combination ofa chemical reaction due to the fluorides and physical-chemical processesdue to the organic part of the organic fluorides. This procedure can becorrelated with in vivo caries reduction insofar as reduced enamelsolubility results in reduced caries formation.

The preparations the invention propose-s to use for the care of teeth,apart from the reagents that have been described, may contain theconventional additives such as polishing agents, e.g., calciumphosphates, calcium carbonate, magnesium carbonate, calcium sulfate, andinsoluble precipitated alkaline earth fluorides. However, to insure theavailability of the fluoride ions over long periods of time, toothpasteformulations should preferably contain calcium polyphosphates, calcitunmetaphosphate, calcium oxyapatite, betaand al-pha-tricalciumorthophosphate. They may also contain surface-active substances whichserve as foam-forming and wetting agents, as Well as aromas, andflavors. In paste-like preparations, mucilages such as tragacanth,carragheen, methyl Example XI A toothpaste is prepared having thefollowing composition:

Percent by weight Sodium carboxymethylcellulose 1.10 Magnesium aluminumsilicate 0.40 Humectants 30.00 Calcium metaphosphate 40.00 Betainepotassium fluoride 2.77 Lauryl alcohol sulfate .70 Flavor and sweeteningQ.s. Water Balance The betaine potassium fluoride employed in the aboveformulation can be replaced with a quantity of cystine hydrofluoride,histidine dihydrofluoride or the mixture of amino acid hydrofluoride ofExample IX sufficient to provide an equivalent amount of fluorinewithout substantially affecting the anticaries efficacy of thecomposition.

Example XII A tooth powder is prepared having the following composition:

Percent by weight Dicalcium phosphate 74.95

Colloidal kaolin 20.0 Aroma substances 1.5 Sodium lauryl sulfonate 3.0Sarcosine-monohydrofluoride 0.55

This composition provides an effective means for caries prophylaxis whenused in the conventional manner. The sarcosine-monohydrofluoride can bereplcaed with alanine hydrofluoride, methionine hydrofluoride orarginine hydrofluoride in an amount sufficient to provide an equalamount of fluorine with substantially equivalent results.

Example XIII A mouthwash is prepared acocrding to the following formula:

Percent by weight Ethyl alcohol 50% 74.0

Aroma substances 5.0 Sulforicinoleate of sodium 7.5 Lysine-potassiumfluoride 13.5

For use, the preparation is strongly diluted, for instance 2 cc. beingmade up to 50 cc. with water, so that the concentration of fluorine inuse is 0.05%.

Example XIV A chewing gum is prepared having the following composition:

Gum base percent by weight 21.0 Estergum parts 30 Coumarone resin do 45Latex (dry) do 15 Paraflin wax (M.P. F.) do 10 Sugar percent by weight"59.5

Corn syrup (Baum 45) do 18.2

Flavoring do Q.s.

Cystine hydrofluoride do 1.0

7 Example XV A fluorine tablet can be prepared according to thefollowing recipe:

In each of the compositions of the above examples the organic fluorideemployed may be replaced by any of the other organic fluorides describedherein.

What is claimed is:

1. An anticariogenic addition compound having the formula R.nX where Ris a lower aliphatic amino acid and X is a soluble fluorine-containingcompound which normally dissociates to yield fluorine ions in aqueoussolution and is selected from the group consisting of HF, alkali metalfluoride, SnF and ZrF, and wherein n is 1 or 2 when X is HF and being 1when X is alkali metal fluoride, SnF, or ZrF 2. An anticariogenicaddition compound according to claim 1 in which X consists of at leastabout 1 mol of HF.

3. An anticariogenic addition compound according to claim 1 in which Xis alkali metal fluoride, SNF, or ZrF 4. A compound according to claim2, namely glycine.HF.

5. A compound according to claim 2, namely, sarcosine.2I-IF.

6. A compound according to claim 2, lysine.HF.

7. A compound according to claim 2, namely, alanine.HF.

8. A compound according to claim 2, namely, betaine.HF.

9. A compound according to claim 3, namely betaine.KF.

10. A compound according to claim 3, namely, betaine.ZrF

11. A compound according to claim 3, namely, glycine.SnF

namely,

12. A compound according to claim 3, namely, 1ysine.KF.

13. A compound according to claim 2, namely, leucine.I-IF.

14. A compound according to claim 2, namely, cystine.HF.

15. A compound according to claim 2, namely cystine amineHF.

16. A compound according to claim 2, namely, sarcosineHF.

17. A compound according to claim 2, namely, methionine.HF.

18. A compound according to claim 2, namely, arginineHF.

19. A compound according to claim 3, namely aspartic acid.SnF

20. A compound according to claim 2, namely, glycine.KF.

References Cited UNITED STATES PATENTS 1,582,472 4/1926 Ikeda 260527 Re.23,344 2/1951 Holgan et a1 260527 XR 2,525,902 10/1950 Holgan et a1.260527 XR 1,634,221 6/1927 Tressler 260-501 1,976,997 10/1934 Kanao260534 2,267,971 12/1941 Braun 260534 2,477,149 7/1949 Sheehan 2605342,579,283 12/1951 Vines 26O534 2,749,213 6/1956 Bruce 260429.7 X3,175,951 3/1965 Tucker 167-93 FOREIGN PATENTS 543,066 7/1957 Canada.

OTHER REFERENCES Greenstein et al., Chemistry of the Amino Acids, JohnWiley & Sons, New York, pp. 649, 650, (1961).

Duschinsky, Chemistry and Industry, January 1934, P. 10.

I Chemical Abstracts, vol. 50, 1956, p. 12819a.

II Chemical Abstracts, vol. 57, 1962, p. 452.

TOBIAS E. LEVOW, Primary Examiner.

H. M. S. SNEED, Assistant Examiner.

